Dirix Carolien, Duvetter Thomas, Loey Ann Van, Hendrickx Marc, Heremans Karel
Department of Chemistry, Faculty of Sciences, Katholieke Universiteit Leuven, Celestijnenlaan 200 D, B-3001 Leuven, Belgium.
Biochem J. 2005 Dec 15;392(Pt 3):565-71. doi: 10.1042/BJ20050721.
The stability of recombinant Aspergillus aculeatus PME (pectin methylesterase), an enzyme with high beta-helix content, was studied as a function of pressure and temperature. The conformational stability was monitored using FTIR (Fourier transform IR) spectroscopy whereas the functional enzyme stability was monitored by inactivation studies. Protein unfolding followed by amorphous aggregation, which makes the process irreversible, was observed at temperatures above 50 degrees C. This could be correlated to the irreversible enzyme inactivation observed at that temperature. Hydrostatic pressure greater than 1 GPa was necessary to induce changes in the enzyme's secondary structure. No enzyme inactivation was observed at up to 700 MPa. Pressure increased PME stability towards thermal denaturation. At 200 MPa, temperatures above 60 degrees C were necessary to cause significant PME unfolding and loss of activity. These results may be relevant for an understanding of the extreme stability of amyloid fibrils for which beta-helices have been proposed as a structural element.
研究了具有高β-螺旋含量的重组棘孢曲霉果胶甲酯酶（PME）的稳定性与压力和温度的关系。使用傅里叶变换红外光谱（FTIR）监测构象稳定性，而通过失活研究监测功能性酶的稳定性。在高于50摄氏度的温度下观察到蛋白质展开并随后发生无定形聚集，这使得该过程不可逆。这可能与在该温度下观察到的酶不可逆失活相关。大于1 GPa的静水压力是诱导酶二级结构变化所必需的。在高达700 MPa时未观察到酶失活。压力增加了PME对热变性的稳定性。在200 MPa时，需要高于60摄氏度的温度才能导致PME显著展开并丧失活性。这些结果可能有助于理解淀粉样原纤维的极端稳定性，其中β-螺旋被认为是一种结构元件。
